ABSTRACT SUMMARY Next-generation sequencing (NGS) has revolutionized the world of genetics, genomics and biomedical research, catalyzing the integration of genome technologies with other biological research fields. The most widely employed NGS is Illumina?s SBS sequencing technology, that generates short read sequences (<200 bp). The Illumina short-read platform alone is insufficient for comprehensive genome analysis, however, as many ?difficult? regions of the genome require additional data to resolve their primary sequence. Third-generation sequencing technologies have now been developed, based on single-molecule real-time sequencing, to overcome this issue and to provide a complementary research tool to NGS technologies. The Pacific Biosciences Sequel II DNA sequencing instrument, for which support is requested in this proposal, is the premier instrument for generating accurate long DNA sequence reads. The Sequel II system, released in April, 2019, accommodates a SMRT Cell with 8 million ZMWs, generates 8x more sequence data than the previous generation Sequel instrument, resulting in a much higher throughput and lower price per base pair. This new instrument will be installed and administrated by the Baylor College of Medicine Human Genome Sequencing Center (BCM-HGSC) and will be made available to NIH supported investigators across BCM. As this will be the first Sequel II instrument at the Texas Medical Center (TMC) we will also enable access by qualified investigators at neighboring institutions. We have identified 9 major users and 3 minor users whose research projects are geared to take advantage of this instrument. We anticipate that the availability of long-read DNA sequencing reads will have a major impact on a multitude of genome studies, providing comprehensive views of genomes and transcriptomes. The participating PIs are all engaged in both basic and clinical research and so this instrument will have direct, broad, and immediate impact on many biological and clinically endeavors. To further enable the instrument users, we will establish a ?user forum? with regular meetings to discuss applications including: whole genome sequencing; variant detection; genome assemblies; targeted sequencing including capture and amplicons; transcriptome sequencing of tissues and single cells; and epigenetic sequencing.